The program of differentiation in human esophageal epithelium was found to be similar to, yet unique from, that of the human epidermis. With the exception of the absence of high molecular weight keratins, the pattern of keratin proteins in human esophageal cells was found to be qualitatively similar to that of epidermal cells. There were, however, dramatic quantitative differences in the array of keratin species between the two tissues. During in vitro cultivation, epithelial cells from these two tissues were found to maintain a differentiative program remarkably similar to that of the native epithelium. Malignant transformation of esophageal epithelial cells was found to be accompanied by significant changes in the array of keratins and in the proportion of cells making cross-linked envelopes. Numerous morphological and biochemical differences have been observed between normal and malignant esophageal epithelial cells in culture. Analysis of keratin proteins extracted from a wide variety of tumors indicates that these proteins may prove to be a valuable adjunct in diagnostic pathology.